Paeoniflorin as a natural mixed-type CYP1B1 inhibitor: enzyme kinetics, molecular simulation and drug-likeness assessment

Paeoniflorin (PF), a monoterpene glycoside from <i>Paeonia</i> species, is known for its antioxidant and anti-inflammatory properties. This study investigated its inhibitory activity against cytochrome P450 1B1 (CYP1B1), a procarcinogen-activating enzyme overexpressed in multiple tumours. Fluorescence-based assays showed dose-dependent inhibition of CYP1B1 by PF, with an IC₅₀ of 0.83 ± 0.17 mM. Kinetic analysis revealed a mixed-type mechanism, suggesting PF binds both free enzyme and enzyme-substrate complex. Molecular docking indicated interactions with active site residues, supported by 100 ns molecular dynamics simulations confirming complex stability. ADMET predictions suggested low toxicity and favourable drug-likeness, though limited oral bioavailability. Collectively, PF is identified as a selective, natural CYP1B1 inhibitor, offering potential for cancer chemoprevention. These findings provide mechanistic insight into CYP1B1 inhibition by monoterpenoids and support future structure-activity relationship (SAR) development.